The use of small interfering RNA molecules (siRNA) is a potent new technology to silence genes and consequently their gene products. It has been reported that RNAi silences genes 10-fold more efficiently than antisense RNA alone. (Rocheleau C E, et al. Wnt signaling and an APC-related gene specify endoderm in early C. elegans embryos. Cell 1997; 90:707-716.) siRNAs have been used to study the role of proteins in signal transduction pathways and it has also been suggested that these molecules might be useful in treating a variety of diseases in which the causative protein is overexpressed. (Arenz C, Schepers U., RNA interference: from an ancient mechanism to a state of the art therapeutic application? Naturwissenschaften 2003; 90:345-359.; Coburn G A, Cullen B R. siRNAs: a new wave of RNA-based therapeutics. J Antimicrob Chemother 2003; 51:753-756.) To avoid nonspecific gene silencing induced by longer double-stranded RNA, small interfering RNAs, a duplex of 21-23 nucleotides, have been used as mediators to degrade target mRNA. (Fire A, Xu S, Montgomery M K, Kostas S A, Driver S E, Mello C C. Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature 1998; 391:806-811.) Once inside the cell, siRNA is incorporated into an RNA-induced silence complex (RISC), a protein-RNA complex that results in unwinding and strand separation of the RNA duplex. The antisense RNA then guides the activated RISC to anneal and cleave the target mRNA. (Hammond S M, Bernstein E, Beach D, Hannon G J. An RNA-directed nuclease mediates post-transcriptional gene silencing in Drosophila cells. Nature 2000; 404:293-296; Reynolds A, Leake D, Boese Q, Scaringe S, Marshall W S, Khvorova A. Rational siRNA design for RNA interference. Nat Biotechnol 2004; 22:326-330; Hammond S M, Boettcher S, Caudy A A, Kobayashi R, Hannon G J. Argonaute2, a link between genetic and biochemical analyses of RNAi. Science 2001; 293:1146-1150; Bernstein E, Caudy A A, Hammond S M, Hannon G J. Role for a bidentate ribonuclease in the initiation step of RNA interference. Nature 2001; 409:363-366.)
Both viral and nonviral carriers have been used to carry siRNA to their cytosolic mRNA target. (Simeoni F, Morris M C, Heitz F, Divita G. Insight into the mechanism of the peptide-based gene delivery system MPG: implications for delivery of siRNA into mammalian cells. Nucleic Acids Res 2003; 31:2717-2724.) To date, however, few peptide carriers have been developed that have proved effective for efficient siRNA delivery to eukaryotic cells (i.e., transfection).
There is a need in the art for pharmaceutical agent delivery systems having transfection efficiencies sufficient to deliver therapeutically effective amounts of siRNA into target cells. In particular, there is a need in the art for improved delivery systems capable of delivering siRNA into the interior of cells. There is also a need in the art for carriers that are stable in serum for delivery systems to be effective both in vitro and in vivo.